微囊化转基因细胞CHO移植抑制血管生长的抗肿瘤治疗

Microencapsulation of recombinant cells is a novel alternative approach to gene therapy of tumors. It is necessary to prepare plenty of microcapsules with better cell viability and higher endostatin production in order to bring this technology into clinic. However, the clinical application of microencapsulation technology is affected because the preparation and culture on large scale is immature. In this paper, we studied the feasibility of microencapsulated recombinant cells treating tumor and large scale preparation of microencapsulated cells. The microencapsulated recombinant CHO cells can grow and express endostatin in culture in vitro and in vivo, and the recombinant endostain secreted from microencapsulated CHO cells can potently inhibit bovine aortic endothelial cells proliferation in vitro (Fig.1) and chicken chorioallantoic membrane angiogenesis in vivo(Fig.2). The results in this study showed that the microcapsules with solid-core, 200μm in diameter and 4-day-culture in vitro were fit for the transplantation of microencapsulated recombinant cells. The peritoneal administration of the encapsulated CHO-endo cells potently inhibited the B16 melanoma growth by 66.4%, decreased the neovascularization of tumor tissue by 59.4%, and increased the survival of the treated mice by 40% after 27 days of treatment. The recombinant CHO-endo cells could expand rapidly in the bioreactor in three dimensions, and the expanded cells could be encapsulated into the microcapsule. The exponential growth phase of recombinant CHO cells (the survival of cells exceed over 90%) with the seeding density of 1-2×106 cells/mL microcapsules benefited to the cells growth and endostatin production during the cell encapsulation procedure, and the time of the CHO-endo retaining in alginate and calcium chloride should be within 3 hours to maintain the cell viability.The cryopreservation is a very good preservation method of recombinant cells. The growth of microencapsulated recombinant CHO cells, the expression of endostatin and the in vivo stability of microcapsule do not change significantly after cryopreservation. However, the 4-day-culture in vitro is necessary to maintain the viability of recombinant cells. The high viable cell density and endostatin production were acquired when the microcapsule percentage was 5% in the culture of the microencapsulated cells, and microencapsulated recombinant cells can be cultured in large scale using B. Braun bioreactor